pGAL
Many different plasmids serve as useful tools in molecular biology. One example is the pGAL plasmid which contains 6751 base pairs and has been cleverly modified by genetic engineering. In the cell, it does not integrate into the bacterial chromosome, but replicates autonomously.
The pGAL plasmid contains the E. coli gene which codes for β-galactosidase. Since the host E. coli does not contain a β-galactosidase gene, only cells transformed by the pGAL plasmid will produce the functional β‑galactosidase enzyme. In the presence of artificial galactosides such as 5-Bromo-4 Chloro 3-indolyl-β-D-galactoside (X-Gal), pGAL colonies appear blue when X-Gal is cleaved by β-galactosidase and forms a colored product.
In other words, cells that express β-galactosidase will cleave X-Gal and the pGAL transformed colonies will be blue.
In addition to the expression and cleavage of X-Gal by β-galactosidase, transformation by pGAL is also demonstrated by resistance to ampicillin. E. coli host cells used in this experiment are not naturally resistant to ampicillin. The plasmid pGAL contains the gene which encodes for β-lactamase that inactivates ampicillin. E. coli cells transformed by pGAL will express the resistance gene product β-lactamase as an extracellular enzyme excreted from E. coli cells. Once outside the cell, the enzyme diffuses into the surrounding medium and inactivates ampicillin.
With time, small "satellite" colonies may appear around a large blue colony. Cells in the small "satellite" or "feeder" colonies are not resistant to ampicillin and have not been transformed with the pGAL plasmid. They are simply growing in a region of agar where β-lactamase has diffused and inactivated the antibiotic ampicillin. The number of satellite colonies increases if the concentration of ampicillin is low or the plates have incubated for longer times.
